An Fmoc Compatible, O to S Shift-Mediated Procedure for the Preparation of C-Terminal Thioester Peptides

Abstract

We report a practical 2-hydroxy-3-mercapto-propionic acid (Hmp)/2-methylpiperidine (2-MP) based Fmoc chemistry procedure to prepare the C-terminal Hmp peptides, which serve as the precursors of C-terminal thioester peptides. The subsequent O to S acyl shift and thiol-exchange mediated thioester conversion of the crude precursor peptides can be accomplished smoothly under mild conditions to provide the desired thioester peptides with good yield and high quality. This is a highly adaptable approach, and we envision its broad application in the preparation of C-terminal thioester peptides.

Chemical synthesis of membrane proteins: a model study on the influenza virus B proton channel

“…As described above, the Hmp unit was attached to a polylysine and/or mini-PEG solubilizing tag and incorporated into the model peptide fragments 1–4 . More precisely, the thioester-forming peptides 2–4 were composed of four consecutive units: the peptide sequence, the Hmp unit, 49 , 51 a spacer (if necessary), and the solubilizing tag ( Fig. 1b ).…”

“…However, under standard NCL conditions, high amounts (20–25%) of the hydrolyzed Cα-carboxyester by-product were reported. 49 , 51 Moreover, the Hmp unit was found to be labile during standard Fmoc-SPPS, which might explain why this elegant strategy has not found a wide use. In 2013, Liu et al observed that the use of 2-methylpiperidine instead of piperidine for the Fmoc-deprotection step nicely solved the instability problem during the Fmoc-SPPS of peptide-oxoesters.…”

“…In 2013, Liu et al observed that the use of 2-methylpiperidine instead of piperidine for the Fmoc-deprotection step nicely solved the instability problem during the Fmoc-SPPS of peptide-oxoesters. 51 …”

Sortase-mediated backbone cyclization of proteins and peptides

“…Several strategies have been employed to overcome this incompatibility: (i) use of the non-nucleophilic bicyclic base 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) instead of piperidine during the Fmoc removal (Clippingdale et al, 2000); (ii) masking of the thioester group by introducing a stable group at the C-terminus which after completion of the whole sequence is converted into a thioester (Brask et al, 2003); and (iii) the use of alternative resin linkers that produce thioesters during acidic work-up after cleavage from the resin (Ingenito et al, 1999). The recent advances in masking strategies (Liu and Mayer, 2013;Zheng et al, 2013) and the use of dedicated resin-linkers (Hou et al, 2010;Ollivier et al, 2010Ollivier et al, , 2014Hemu et al, 2013;Taichi et al, 2013) have made peptides with a C-terminal thioester group readily accessible for Fmoc-chemistry.…”

Intramolecular acyl transfer in peptide and protein ligation and synthesis

“…These challenges led to the development of many N‐to‐S and also O‐to‐S acyl transfer strategies to obtain peptide thioesters. Peptide thioesters have been successfully synthesized by O‐to‐S acyl transfer methods with C‐terminal mercaptoethanol ester , phenolic esters and 2‐hydroxy‐mercaptopropanoate ester . There are also reports that utilize this reaction for in situ or one‐pot formation of thioesters to achieve NCL due to the fast kinetics of O‐to‐S acyl transfer compared with N‐to‐S acyl transfer ; S‐to‐O migration has been observed via a special silatropic ‘switch’ where thiol acids were activated to form S‐silyl ester.…”